Cellular Reproduction CHAPTER 14 Cellular Reproduction

Overview of the cell cycle DNA replication DNA Damage and repair Nuclear and Cell Division Regulation of the Cell cycle Growth Factors and Cell Proliferation WILL NOT BE Included in the exam

Introduction Cells reproduce by the process of cell division. Mitosis leads to cells that are genetically identical to their parent. Meiosis leads to production of cells with half of the genetic content of the parent.

14.1 The Cell Cycle (1) The cell cycle is the series of stages that a cell goes through. It consists of the M phase and the interphase. The M phase includes the process of mitosis and cytokinesis. Mitosis last about an hour or so. Interphase constitutes the majority of the cell cycle and lasts longer than the M phase (~23 hrs); it may extend for days, week, or longer.

An overview of the eukaryotic cell cycle

The Eukaryotic Cell Cycle Cell growth === Cell division=== Cell proliferation -Duplication of DNA -Replication of DNA -DNA synthesis ---Mitosis/ Cytokinesis ---Regulation of Cell cycle Cell cycle Mitosis ~0.5 hrs 1-Starts—de/condensation- chromatin 2a-sister chromatids 2b-nuclear envelop disorganize 2c-mitotic spindle 3-cytokinesis G1 8 to 10 hrs S 6 to 8 hrs G2 4 to 6 hrs Go: the cell is out the cell cycle Terminal differentiation Go -Protein synthesis ? ~23.5 hrs

The Cell Cycle (2) Interphase includes G1, S, and G2 periods. G1 takes place between the end of mitosis and the beginning of DNA replication. DNA replication occurs during the S phase. G2 occurs between the end of S and the beginning of mitosis. Incorporation of 3H-thymidine 3H 1-Add 2-Wash (Pulse and chase)

The Cell Cycle (3) Cell cycles in vivo Three cell types are distinguished based on their capacity to grow and divide. Nerve cells have lost the ability to divide. Liver cells divides in function of stimulus Stem cells will divide frequently. Stem cells have asymmetric cell division in which the daughter cells have different fates.

The Cell Cycle (4) Control of the Cell Cycle Cell cycle is focused on initiation of DNA replication and initiation of mitosis. The cytoplasm contains factors that regulate the state of the nucleus. The cytoplasm of a replicating cell contains factors that stimulate initiation of DNA synthesis. The cytoplasm of a cell undergoing mitosis contains factors that trigger chromosomal condensation.

The stages of mitosis in animal and plants cells

The Phases of Mitosis in an Animal Cell What kind of Signal? G1 to S Chromatin begins “condensation Centrosome duplicates (S phase)

The Phases of Mitosis in an Animal Cell Discrete chromosomes visible under microscope Nucleus is not clear Nucleolus ~disappears Centrosomes move to opposite sides of nucleus (MTOC, Aster, centrioles) Mitotic spindle assembles (microtubules)

The Phases of Mitosis in an Animal Cell Prometaphase=Late prophase Nuclear envelope fragmentation Centrosomes move to opposite sides of nucleus Spindle MTs make contact with chromosomes ---centromere (sequence repeated DNA sequence=heterochromatin =CEN sequences) ---kinetochore (protein-like containing structure)

Three types of spindle microtubules (MT) Polar microtubules Kinetochore microtubules

The Phases of Mitosis in an Animal Cell Perfect and equidistant distribution of the chromosomes. Metaphase plate Spindle perturbing drugs arrest cells in metaphase Colchicine and Nocodazole’s effect

Attachment of Chromosomes to the Mitotic Spindle CEN proteins

The Phases of Mitosis in an Animal Cell Anaphase= separation of sister chromatids Anaphase A (early) – chromosomes pulled toward spindle poles Anaphase B (late) – poles move away from each other

Kinetochore MT= shorter The Two Types of Movement Involved in Chromosome Separation During Anaphase Kinetochore MT= shorter Spindle poles= move away

The Phases of Mitosis in an Animal Cell Reorganization= new cell

Microtubule Polarity in the Mitotic Spindle Polarity of MT

Microtubule Polarity in the Mitotic Spindle Chromosome alignment and separation Separated by microtubules, also requires additional proteins like Topoisomerase II, changes in adhesive proteins, (anaphase) and motor proteins (three different roles--mitotic motors-)

Mitotic Motors Kinetochore microtubules Plus & minus ends Depolymerases binds MT and induces depolymerization (chromokinesin, C-terminal and bipolar kinesin) Polar microtubules ATP hydrolysis is require for MT sliding (C-terminal and bipolar kinesin) Astral microtubules MT associated (cytoplasmic dynein) with cell cortex at plasma membrane (actin microfilaments) Taxol: it blocks MT depolymerization

Cytokinesis in an Animal Cell Cleavage furrow associated with the contractile ring (Actin and Myosin). Mechanism: ? De-phosphorylation and phosphorylation Rho small GTPases

Cytokinesis and Cell Plate Formation in a Plant Cell Vesicles from Golgi and ER Youtube====50-year cell division puzzle solved

2 3 1 Typical eukaryotic cell cycle --- G1, S, G2, M ---Control / Regulation Systems: 1--Appropriate time and sequence of events 2--Each event must be completed 3--Respond to external conditions (nutrients and growth factors) Cell cycle control: Key Transition Points in the Cell Cycle Chromosome segregation 2 3 1

Cell Fusion Evidence for the Role of Cytoplasmic Chemical Signals in Cell Cycle Regulation Specific molecules present in the cytoplasm that induce the transformation from S to G1 (DNA replication genes=proteins) from M to G1 (cell cycle regulation Gene: cdc2=cell division cycle --regulate the transition from G2 to M --cdc2: protein kinase Regulation by phosphorylation?

Experimental demonstration that cells contain factors that stimulate entry into mitosis

How is the cell cycle regulated? Cell Cycle Regulation Kinase Phosphatase Substrate (P2) Substrate (P1) (Cyclins) =substrate (Cyclin-dependent kinase) -CDK- Mitotic M-cyclins Mitotic M-cdks G1 cyclins G1 cdks S cyclins Cdc2 (Cell Division Cycle ) =kinase Cdc=cdk http://www-rcf.usc.edu/~forsburg/cclecture.html

Evidence for the Existence of MPF Maturation promoting factor=MPF =mitotic cdk-cyclin complex ----control the G2 to M transition

Fluctuating Levels of Mitotic Cyclin and MPF During the Cell Cycle Maximal Enzymatic Activity during the G2-M transition Substrate:? Regulation:? CDK Relative activity Mitotic cyclin CDK

mitotic cdk-cyclin complex=MPF Phosphorylation and Dephosphorylation in the Activation of a Cdk-Cyclin Complex Ser= variable Thr= 161 Tyr=15 MPF-i MPF-a mitotic cdk-cyclin complex=MPF

Fluctuating Levels of Mitotic Cyclin and MPF During the Cell Cycle Maximal Enzymatic Activity during the G2-M transition Substrate:? Regulation:? Relative activity CDK

The Mitotic Cdk Cycle Cell Cycle Control-I 1-Phosphorylation of lamin in the nuclear lamina induces depolarization 2-Phosphorylation of condensin induces chromosome condensation 3-Phosphorylation of MT induces polymerization 4-Phosphorylation of Anaphase promoting degradation complex. -APC-(ubiquitin ligase) Cell Cycle Control-I

The Anaphase-Promoting Complex and the Spindle Checkpoint Mitotic CDK-cyclin Anaphase promoting complex (ubiquitin ligase)

The Anaphase-Promoting Complex and the Spindle Checkpoint Regulation of the anaphase promoting complex: Binding with cdc20 APC cdc20

-degradate mitotic cyclins complex APC cdh1 -degradate mitotic cyclins complex SCF complex SCF controls G1/S through G2/M transitions p27 and p21 (cyclin-dependent-kinase inhibitors (CKIs) G1/S cyclin, Cyclin E

Role of the Rb (Retinoblastoma) Protein in Cell Cycle Control G1 cdk cyclin Cell Cycle Control-II

How is the cell cycle regulated? Cell Cycle Regulation Mitotic M-cyclins Mitotic M-cdks /A G1 cyclins G1 cdks S cyclins , 6

DNA Damage: p53-dependent and independent pathways can not be repaired-- Cell Cycle Control-III ATM kinase=ataxia telangiectasia mutated/mutation ATM=Ionizing radiation

DNA Damage: p53-dependent and independent pathways ATR= UV radiation ATR kinase: Inactivation of cyclin

http://www.epa.gov

Checkpoints=Cell Cycle Control Spindle checkpoint Kinetochore attachment Mad, Bub, Cdc20 DNA replication checkpoint Completion of DNA synthesis G2/M transition Phosphorylation of mitotic cyclin DNA damage checkpoint Cell cycle halted at various stages p53 – guardian of the genome A. DNA integrity (DNA damage) Timer Cdk Cyclins B. Processes (complete)

General Model for Cell Cycle Regulation APC complex Regulation Nutrients Mitogens EGF PDGF Growth factors Extra-cellular signals APC / SCF complex

Growth Factor Signaling via the Ras Pathway -bacteria and yeast cell proliferation/division Nutrients Glucose -mammalian cells cell proliferation/division Growth factor = mitogens

Growth Factor Signaling via the Ras Pathway

Growth Factor Signaling via the Ras Pathway PM or endosomes Mitogenic Activated Protein Raf=MAPkkk MEK=MAPkk ERK=MAPk Tyrosine Serine/Threonine

The PI3K-Akt Signaling Pathway Insulin (I) and Insulin-like growth factor (IGF)

Cell Growth / Proliferation EGF/ PDGF TGF- (+) (-) Cell Growth / Proliferation Phosphorylation of Smad Activation of p27, p21 and p15 (Cdk inhibitors) MAPK Mothers Against Decapentaplegic (MAD) Small Mothers Against paralysis (SMA)

General Model for Cell Cycle Regulation APC complex Regulation Nutrients Mitogens (+) (-) TGF- EGF PDGF (+) Growth factors Extra-cellular signals APC / SCF complex